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Abstract

Arachidonic acid (ARA) metabolites produced by cyclo-oxygenase and lipoxygenase are important mediators maintaining physiological renal function. However, the effects of exogenous ARA on kidney function in vivo remain unknown. This study examined the effects of long-term oral ARA administration on normal renal function as well as inflammation and oxidative stress in aged rats. In addition, we measured levels of renal eicosanoids and docosanoids using liquid chromatography–tandem mass spectrometry. Control or ARA oil (240 mg/kg body weight/day) was orally administered to 21-month-old Wistar rats for 13 weeks. Levels of plasma creatinine, blood urea nitrogen, inflammatory and anti-inflammatory cytokines, reactive oxygen species, and lipid peroxidation were not significantly different between the two groups. The ARA concentration in the plasma, kidney, and liver increased in the ARA-administered group. In addition, levels of free-form ARA, prostaglandin E2, and 12- and 15-hydroxyeicosatetraenoic acid increased in the ARA-administered group, whereas renal concentration of docosahexaenoic acid and eicosapentaenoic acid decreased in the ARA-administered group. Levels of docosahexaenoic acid-derived protectin D1, eicosapentaenoic acid-derived 5-, and 18-hydroxyeicosapentaenoic acids, and resolvin E2 and E3 decreased in the ARA-administered group. Our results indicate that long-term ARA administration led to no serious adverse reactions under normal conditions and to a decrease in anti-inflammatory docosahexaenoic acid- and eicosapentaenoic acid-derived metabolites in the kidneys of aged rats. These results indicate that there is a possibility of ARA administration having a reducing anti-inflammatory effect on the kidney.

Animals and treatments

Rats (Jcl: Wistar) purchased from CLEA Japan (Osaka, Japan) were housed in a room under controlled temperature (23 ± 2°C), humidity (50 ± 10%), and light–dark cycle (light: 07:00–19:00; dark: 19:00–07:00). They were fed a fish oil-deficient diet (F-1™ Funabashi Farm, Funabashi, Japan) and water ad libitum. Inbred second-generation male rats, fed the same F-1 diet, were used for our study. Fatty acid composition in the F-1 diet was described previously []. ARA oil was obtained from Cargill Alking Bioengineering (Wuhan and Hubei, China) [,]. The fatty acid compositions of the ARA and control oils are shown in . Twenty one-month-old male rats were divided into two groups: the control group was orally administrated with the control oil and the ARA group was administrated ARA oil (240 mg ARA/kg body weight/day) for 13 weeks. After a 16-h fast, rats were anesthetized with intraperitoneal sodium pentobarbital (65 mg/kg body weight), following which their plasma, kidneys, and liver were removed, immediately frozen in liquid nitrogen, and stored at −30°C until further use. The kidneys and liver were homogenized in phosphate buffer (pH, 7.4) using a Teflon homogenizer (AGC techno glass Co., Ltd. Shizuoka, Japan). The homogenates were immediately frozen in liquid nitrogen and stored at −30°C until use. Concentrations of creatinine and blood urea nitrogen (BUN) were determined in plasma samples using an automatic analyzer (BiOLiS 24i; Tokyo Boeki Medical System Ltd., Tokyo, Japan).

Levels of eicosanoids and docosanoids in the kidney

Kidney analyses revealed an increase in renal formation of PGE2, 12-HETE, and 15-HETE in the ARA-administered group (). Moreover, endogenous formation of DHA-derived PD1 and EPA-derived 5-HEPE, 18-HEPE, RvE2, and RvE3 decreased significantly in the ARA-administered group. Nonesterified ARA levels in the kidney increased significantly in the ARA-administered group, whereas levels of nonesterified EPA and DHA were not significantly different between the two groups.

Discussion

Concentrations of non-esterified EPA and DHA increased slightly, but not significantly, in the ARA-administered group compared with those in the control group, although total EPA and DHA concentrations decreased in the kidney. Levels of EPA-derived eicosanoids 5-HEPE, 18-HEPE, RvE2, and RvE3 and the DHA-derived docosanoids PD1 decreased significantly in the ARA-administered group. These results suggest that ARA directly competes with the storage of EPA and DHA at the sn-2 position in phospholipids and blocks the production of EPA- and DHA-derived metabolites []. We have reported that the DHA-derived docosanoids RvDs and PD1 protect renal damage progression induced by metabolic syndrome []. EPA-derived eicosanoids and DHA-derived docosanoids are endogenous mediators with potent anti-inflammatory actions in the kidneys []. Administration of RvDs or PD1 to mice prior to ischemia results in a reduction in functional and morphological renal injury []. Hong and Lu demonstrated that RvDs and PD1 repress renal interstitial fibrosis, and PD1 inhibits the inflammatory response and promotes renoprotective heme-oxygenase-1 expression during acute kidney injury []. Heme-oxygenase-1 decreases acute tubular necrosis and significantly reduces COX-2 and microsomal PGE synthase expression []. Our study demonstrated that RvE2, RvE3, and PD1 levels decreased significantly in the ARA-administered group, indicating that anti-inflammatory defense may be attenuated by ARA administration.